1. Field of the Invention
One of the most important constraints on the yields of food and cash crops worldwide can be attributed to insect attack. Based on 1987 figures, approximately 37% of all crops produced worldwide are lost to pests such as insects (13%), disease (12%), and weeds and grasses (12%). Annually, large sums of money are spent on chemical pesticides to reduce these levels of crop damage. In 1987 the insecticide expenditures for the three crops receiving the highest insecticide input--cotton, maize and rice--was almost $4000 million. Approximately $3000 million was spent on purchasing the insecticide and $1000 million for application. In the U.S. alone, over $400 million is spent each year for control of lepidopterans.
Dependance on chemical pesticides for crop damage control is not only expensive, but it is also detrimental to the environment and unhealthy for the animal population. Many chemical insecticides, particularly organophosphates and carbamates, are neurotoxic to a wide range of animals from honey bees to humans. A number of them have been discontinued because of their toxic properties. Thus, scientists are currently seeking alternatives to the conventional chemical approach to crop pest management, and one approach is the investigation of plant-mediated, and thus more environmentally friendly, methods and products.
Scientists have long used cross-breeding and hybridization techniques to provide plants having particular desired traits such as increased hardiness, nutritional value, taste, appearance, etc., but these techniques are at best lengthy, time-consuming processes which do not necessarily result in the achievement of a particular goal. The advent of genetic engineering, however, provided the opportunity to introduce genetic material directly into a plant, which, upon expression in the plant, would result in a desired effect.
2. Description of the Prior Art
A limited number of insect-control agents are currently available for genetic engineering into plants. The protein delta endotoxins from the microorganism Bacillus thuringiensis (Bt) have been the most widely studied in transformed plants, and the class of proteins known as proteinase inhibitors, when present at relatively high levels in the diet, has been shown to be effective against certain insects. The potential disadvantages to using transgenic Bt plants are that effective concentrations may be difficult to achieve in the plant and that insect resistance may develop with time. The high levels of protein required for insect killing and the potential need to target protein expression to specific plant organs are problems associated with usage of proteinase inhibitors as insect-control agents.
Phytohormones are known to have pivotal roles in promoting normal growth and development of plants and may also contribute to the mechanisms of defense (Gatehouse, 1991; Nicholson, 1992). Cytokinins are among the most active plant substances discovered and have been implicated in the physiological and biochemical processes with marked effects on flowering, fruit set and ripening, leaf senescence, seed germination and stomatal function. Exogenously applied cytokinins have been shown to suppress the induction of hypersensitive necrosis by viruses (Bailiss et al., 1977; Balazs and Kiraly, 1981). High endogenous cytokinin levels in non-rooting tobacco shoot lines (T-cyt) transformed with a gene involved in cytokinin biosynthesis caused an increase in the expression of defense-related mRNAs (Memelink et al., 1987). A group of pathogenesis related proteins encoded by these genes is coordinately induced by wounding and pathogenic infections (Chen and Varner, 1985; Ward et al., 1991).
Cytokinins as well as other plant hormones have commercial applications as bioregulators and, in combination with endogenous hormones, may protect plants from pests and pathogens by inducing physiological changes in the plants (Hallahan et al., 1991; Hedin et al., 1988; Thomas and Balkesley, 1987).
Cytokinins have also been shown to influence secondary metabolic pathways whose products exhibit insecticidal properties (Teutonico et al., 1991). Utilization of numerous secondary metabolites in crop protection, either by conventional plant breeding or by genetic engineering, is currently being evaluated (Gatehouse et al., 1992; Hallahan, supra).
Thus, although phytohormones, and cytokinins in particular, have been implicated in conferring resistance to insects in plants, the role of cytokinins in such resistance has not heretofore been evaluated nor have any attempts been made to utilize endogenous cytokinins to confer increased resistance to insects in plants.